Our laboratory has developed a technology to combine proteomic, phosphoproteomic, and genomic biomarkers simultaneously in a clinically relevant and clinically applicable manner. As the Multi-parameter Flow Cytometric Genomics Core, we will be applying our novel multiplexing technology to address the following specific aims. 1) Develop the application of multiparameter flow cytometry gene expression profiling to deconvolute the evolution of the immune response in the peripheral blood to a kidney or heart transplant. We will combine an array of lineage- and activation-specific antibodies that define each cell subset and its state and simultaneous multi-parameter mRNA transcript detection in a single run. 2) Develop the application of multiparameter flow cytometry proteomics to deconvolute the evolution of the immune response in the peripheral blood to a kidney or heart transplant. We will combine the same array of lineage- and activation-specific antibodies that define each cell subset and its state used in Aim 1 and simultaneous multi-parameter protein and/or phosphoprotein detection in a single run. 3) Develop the diagnostic applications of multiparameter flow cytometry done as a function of time following kidney and heart transplantation to identify blood cell subset-specific changes in composition and cell activation during a recognized clinical event or predicting rejection. We will also correlate changes in cell subset composition as a function of time, clinical events and outcomes with the whole blood profiling of gene transcripts, alternative splicing, miRNA expression and phosphoproteomlcs done in parallel on the same clinical samples by Project #1 and Project #2.